HEADER MEMBRANE PROTEIN 15-JAN-25 9LJJ TITLE CRYO-EM STRUCTURE OF A NANOBODY BOUND HELIORHODOPSIN COMPND MOL_ID: 1; COMPND 2 MOLECULE: HELIORHODOPSIN; COMPND 3 CHAIN: A, B; COMPND 4 ENGINEERED: YES; COMPND 5 OTHER_DETAILS: SEQUENCE REFERENCE FOR SOURCE ORGANISM ESCHERICHIA COMPND 6 COLI IS NOT AVAILABLE IN UNIPROT AT THE TIME OF BIOCURATION. CURRENT COMPND 7 SEQUENCE REFERENCE IS FROM UNIPROT ID A0A2R4S913.; COMPND 8 MOL_ID: 2; COMPND 9 MOLECULE: NANOBODY B4; COMPND 10 CHAIN: M, N; COMPND 11 ENGINEERED: YES SOURCE MOL_ID: 1; SOURCE 2 ORGANISM_SCIENTIFIC: ESCHERICHIA COLI; SOURCE 3 ORGANISM_TAXID: 562; SOURCE 4 EXPRESSION_SYSTEM: SPODOPTERA FRUGIPERDA; SOURCE 5 EXPRESSION_SYSTEM_TAXID: 7108; SOURCE 6 MOL_ID: 2; SOURCE 7 ORGANISM_SCIENTIFIC: VICUGNA PACOS; SOURCE 8 ORGANISM_TAXID: 30538; SOURCE 9 EXPRESSION_SYSTEM: ESCHERICHIA COLI; SOURCE 10 EXPRESSION_SYSTEM_TAXID: 562 KEYWDS BACTERIAL RHODOPSIN, MEMBRANE PROTEIN EXPDTA ELECTRON MICROSCOPY AUTHOR Y.HE,R.XIA REVDAT 1 12-FEB-25 9LJJ 0 JRNL AUTH R.XIA,M.SUN,Y.LU,N.WANG,A.ZHANG,C.GUO,Z.XU,X.CAI,Y.HE JRNL TITL CRYO-EM STRUCTURE OF A NANOBODY-BOUND HELIORHODOPSIN. JRNL REF BIOCHEM.BIOPHYS.RES.COMMUN. V. 750 51398 2025 JRNL REFN ESSN 1090-2104 JRNL PMID 39889627 JRNL DOI 10.1016/J.BBRC.2025.151398 REMARK 2 REMARK 2 RESOLUTION. 3.23 ANGSTROMS. REMARK 3 REMARK 3 REFINEMENT. REMARK 3 SOFTWARE PACKAGES : PHENIX REMARK 3 RECONSTRUCTION SCHEMA : NULL REMARK 3 REMARK 3 EM MAP-MODEL FITTING AND REFINEMENT REMARK 3 PDB ENTRY : NULL REMARK 3 REFINEMENT SPACE : NULL REMARK 3 REFINEMENT PROTOCOL : NULL REMARK 3 REFINEMENT TARGET : NULL REMARK 3 OVERALL ANISOTROPIC B VALUE : NULL REMARK 3 REMARK 3 FITTING PROCEDURE : NULL REMARK 3 REMARK 3 EM IMAGE RECONSTRUCTION STATISTICS REMARK 3 NOMINAL PIXEL SIZE (ANGSTROMS) : NULL REMARK 3 ACTUAL PIXEL SIZE (ANGSTROMS) : NULL REMARK 3 EFFECTIVE RESOLUTION (ANGSTROMS) : 3.230 REMARK 3 NUMBER OF PARTICLES : 239778 REMARK 3 CTF CORRECTION METHOD : NONE REMARK 3 REMARK 3 EM RECONSTRUCTION MAGNIFICATION CALIBRATION: NULL REMARK 3 REMARK 3 OTHER DETAILS: NULL REMARK 4 REMARK 4 9LJJ COMPLIES WITH FORMAT V. 3.30, 13-JUL-11 REMARK 100 REMARK 100 THIS ENTRY HAS BEEN PROCESSED BY PDBC ON 19-JAN-25. REMARK 100 THE DEPOSITION ID IS D_1300055773. REMARK 245 REMARK 245 EXPERIMENTAL DETAILS REMARK 245 RECONSTRUCTION METHOD : SINGLE PARTICLE REMARK 245 SPECIMEN TYPE : NULL REMARK 245 REMARK 245 ELECTRON MICROSCOPE SAMPLE REMARK 245 SAMPLE TYPE : PARTICLE REMARK 245 PARTICLE TYPE : POINT REMARK 245 NAME OF SAMPLE : DIMER COMPLEX REMARK 245 SAMPLE CONCENTRATION (MG ML-1) : NULL REMARK 245 SAMPLE SUPPORT DETAILS : NULL REMARK 245 SAMPLE VITRIFICATION DETAILS : NULL REMARK 245 SAMPLE BUFFER : NULL REMARK 245 PH : 7.00 REMARK 245 SAMPLE DETAILS : NULL REMARK 245 REMARK 245 DATA ACQUISITION REMARK 245 DATE OF EXPERIMENT : NULL REMARK 245 NUMBER OF MICROGRAPHS-IMAGES : NULL REMARK 245 TEMPERATURE (KELVIN) : NULL REMARK 245 MICROSCOPE MODEL : FEI TECNAI F30 REMARK 245 DETECTOR TYPE : GATAN K2 SUMMIT (4K X 4K) REMARK 245 MINIMUM DEFOCUS (NM) : 1200.00 REMARK 245 MAXIMUM DEFOCUS (NM) : 2200.00 REMARK 245 MINIMUM TILT ANGLE (DEGREES) : NULL REMARK 245 MAXIMUM TILT ANGLE (DEGREES) : NULL REMARK 245 NOMINAL CS : NULL REMARK 245 IMAGING MODE : BRIGHT FIELD REMARK 245 ELECTRON DOSE (ELECTRONS NM**-2) : 6000.00 REMARK 245 ILLUMINATION MODE : FLOOD BEAM REMARK 245 NOMINAL MAGNIFICATION : NULL REMARK 245 CALIBRATED MAGNIFICATION : NULL REMARK 245 SOURCE : FIELD EMISSION GUN REMARK 245 ACCELERATION VOLTAGE (KV) : 300 REMARK 245 IMAGING DETAILS : NULL REMARK 247 REMARK 247 ELECTRON MICROSCOPY REMARK 247 THE COORDINATES IN THIS ENTRY WERE GENERATED FROM ELECTRON REMARK 247 MICROSCOPY DATA. PROTEIN DATA BANK CONVENTIONS REQUIRE REMARK 247 THAT CRYST1 AND SCALE RECORDS BE INCLUDED, BUT THE VALUES REMARK 247 ON THESE RECORDS ARE MEANINGLESS EXCEPT FOR THE CALCULATION REMARK 247 OF THE STRUCTURE FACTORS. REMARK 300 REMARK 300 BIOMOLECULE: 1 REMARK 300 SEE REMARK 350 FOR THE AUTHOR PROVIDED AND/OR PROGRAM REMARK 300 GENERATED ASSEMBLY INFORMATION FOR THE STRUCTURE IN REMARK 300 THIS ENTRY. THE REMARK MAY ALSO PROVIDE INFORMATION ON REMARK 300 BURIED SURFACE AREA. REMARK 350 REMARK 350 COORDINATES FOR A COMPLETE MULTIMER REPRESENTING THE KNOWN REMARK 350 BIOLOGICALLY SIGNIFICANT OLIGOMERIZATION STATE OF THE REMARK 350 MOLECULE CAN BE GENERATED BY APPLYING BIOMT TRANSFORMATIONS REMARK 350 GIVEN BELOW. BOTH NON-CRYSTALLOGRAPHIC AND REMARK 350 CRYSTALLOGRAPHIC OPERATIONS ARE GIVEN. REMARK 350 REMARK 350 BIOMOLECULE: 1 REMARK 350 AUTHOR DETERMINED BIOLOGICAL UNIT: TETRAMERIC REMARK 350 APPLY THE FOLLOWING TO CHAINS: A, B, M, N REMARK 350 BIOMT1 1 1.000000 0.000000 0.000000 0.00000 REMARK 350 BIOMT2 1 0.000000 1.000000 0.000000 0.00000 REMARK 350 BIOMT3 1 0.000000 0.000000 1.000000 0.00000 REMARK 465 REMARK 465 MISSING RESIDUES REMARK 465 THE FOLLOWING RESIDUES WERE NOT LOCATED IN THE REMARK 465 EXPERIMENT. (M=MODEL NUMBER; RES=RESIDUE NAME; C=CHAIN REMARK 465 IDENTIFIER; SSSEQ=SEQUENCE NUMBER; I=INSERTION CODE.) REMARK 465 REMARK 465 M RES C SSSEQI REMARK 465 MET A 1 REMARK 465 ALA A 2 REMARK 465 LYS A 3 REMARK 465 MET B 1 REMARK 465 ALA B 2 REMARK 465 LYS B 3 REMARK 465 ALA M 112 REMARK 465 ALA M 113 REMARK 465 ALA M 114 REMARK 465 LEU M 115 REMARK 465 GLU M 116 REMARK 465 HIS M 117 REMARK 465 HIS M 118 REMARK 465 HIS M 119 REMARK 465 HIS M 120 REMARK 465 HIS M 121 REMARK 465 HIS M 122 REMARK 465 ALA N 112 REMARK 465 ALA N 113 REMARK 465 ALA N 114 REMARK 465 LEU N 115 REMARK 465 GLU N 116 REMARK 465 HIS N 117 REMARK 465 HIS N 118 REMARK 465 HIS N 119 REMARK 465 HIS N 120 REMARK 465 HIS N 121 REMARK 465 HIS N 122 REMARK 500 REMARK 500 GEOMETRY AND STEREOCHEMISTRY REMARK 500 SUBTOPIC: TORSION ANGLES REMARK 500 REMARK 500 TORSION ANGLES OUTSIDE THE EXPECTED RAMACHANDRAN REGIONS: REMARK 500 (M=MODEL NUMBER; RES=RESIDUE NAME; C=CHAIN IDENTIFIER; REMARK 500 SSEQ=SEQUENCE NUMBER; I=INSERTION CODE). REMARK 500 REMARK 500 STANDARD TABLE: REMARK 500 FORMAT:(10X,I3,1X,A3,1X,A1,I4,A1,4X,F7.2,3X,F7.2) REMARK 500 REMARK 500 EXPECTED VALUES: GJ KLEYWEGT AND TA JONES (1996). PHI/PSI- REMARK 500 CHOLOGY: RAMACHANDRAN REVISITED. STRUCTURE 4, 1395 - 1400 REMARK 500 REMARK 500 M RES CSSEQI PSI PHI REMARK 500 SER A 85 -158.30 -83.28 REMARK 500 ALA A 182 70.97 58.95 REMARK 500 PRO A 192 -177.03 -69.03 REMARK 500 SER B 85 -158.28 -83.28 REMARK 500 ALA B 182 70.97 58.88 REMARK 500 PRO B 192 -177.05 -69.00 REMARK 500 ARG M 45 169.09 179.98 REMARK 500 THR M 52 12.56 50.89 REMARK 500 ASP M 61 -0.67 66.83 REMARK 500 ALA M 74 0.15 57.56 REMARK 500 PRO M 87 45.84 -78.54 REMARK 500 ALA M 91 -173.57 -170.48 REMARK 500 THR M 97 -155.94 -149.11 REMARK 500 ARG N 45 169.07 173.30 REMARK 500 THR N 52 12.50 50.90 REMARK 500 ASP N 61 -0.66 66.74 REMARK 500 PRO N 87 45.79 -78.52 REMARK 500 ALA N 91 -173.64 -170.51 REMARK 500 THR N 97 -155.88 -149.10 REMARK 500 REMARK 500 REMARK: NULL REMARK 900 REMARK 900 RELATED ENTRIES REMARK 900 RELATED ID: EMD-63157 RELATED DB: EMDB REMARK 900 CRYO-EM STRUCTURE OF A NANOBODY BOUND HELIORHODOPSIN DBREF1 9LJJ A 1 256 UNP A0A2R4S913_9ACTN DBREF2 9LJJ A A0A2R4S913 1 256 DBREF1 9LJJ B 1 256 UNP A0A2R4S913_9ACTN DBREF2 9LJJ B A0A2R4S913 1 256 DBREF 9LJJ M 1 122 PDB 9LJJ 9LJJ 1 122 DBREF 9LJJ N 1 122 PDB 9LJJ 9LJJ 1 122 SEQRES 1 A 256 MET ALA LYS PRO THR VAL LYS GLU ILE LYS SER LEU GLN SEQRES 2 A 256 ASN PHE ASN ARG ILE ALA GLY VAL PHE HIS LEU LEU GLN SEQRES 3 A 256 MET LEU ALA VAL LEU ALA LEU ALA ASN ASP PHE ALA LEU SEQRES 4 A 256 PRO MET THR GLY THR TYR LEU ASN GLY PRO PRO GLY THR SEQRES 5 A 256 THR PHE SER ALA PRO VAL VAL ILE LEU GLU THR PRO VAL SEQRES 6 A 256 GLY LEU ALA VAL ALA LEU PHE LEU GLY LEU SER ALA LEU SEQRES 7 A 256 PHE HIS PHE ILE VAL SER SER GLY ASN PHE PHE LYS ARG SEQRES 8 A 256 TYR SER ALA SER LEU MET LYS ASN GLN ASN ILE PHE ARG SEQRES 9 A 256 TRP VAL GLU TYR SER LEU SER SER SER VAL MET ILE VAL SEQRES 10 A 256 LEU ILE ALA GLN ILE CYS GLY ILE ALA ASP ILE VAL ALA SEQRES 11 A 256 LEU LEU ALA ILE PHE GLY VAL ASN ALA SER MET ILE LEU SEQRES 12 A 256 PHE GLY TRP LEU GLN GLU LYS TYR THR GLN PRO LYS ASP SEQRES 13 A 256 GLY ASP LEU LEU PRO PHE TRP PHE GLY CYS ILE ALA GLY SEQRES 14 A 256 ILE VAL PRO TRP ILE GLY LEU LEU ILE TYR VAL ILE ALA SEQRES 15 A 256 PRO GLY SER THR SER ASP VAL ALA VAL PRO GLY PHE VAL SEQRES 16 A 256 TYR GLY ILE ILE ILE SER LEU PHE LEU PHE PHE ASN SER SEQRES 17 A 256 PHE ALA LEU VAL GLN TYR LEU GLN TYR LYS GLY LYS GLY SEQRES 18 A 256 LYS TRP SER ASN TYR LEU ARG GLY GLU ARG ALA TYR ILE SEQRES 19 A 256 VAL LEU SER LEU VAL ALA LYS SER ALA LEU ALA TRP GLN SEQRES 20 A 256 ILE PHE SER GLY THR LEU ILE PRO ALA SEQRES 1 B 256 MET ALA LYS PRO THR VAL LYS GLU ILE LYS SER LEU GLN SEQRES 2 B 256 ASN PHE ASN ARG ILE ALA GLY VAL PHE HIS LEU LEU GLN SEQRES 3 B 256 MET LEU ALA VAL LEU ALA LEU ALA ASN ASP PHE ALA LEU SEQRES 4 B 256 PRO MET THR GLY THR TYR LEU ASN GLY PRO PRO GLY THR SEQRES 5 B 256 THR PHE SER ALA PRO VAL VAL ILE LEU GLU THR PRO VAL SEQRES 6 B 256 GLY LEU ALA VAL ALA LEU PHE LEU GLY LEU SER ALA LEU SEQRES 7 B 256 PHE HIS PHE ILE VAL SER SER GLY ASN PHE PHE LYS ARG SEQRES 8 B 256 TYR SER ALA SER LEU MET LYS ASN GLN ASN ILE PHE ARG SEQRES 9 B 256 TRP VAL GLU TYR SER LEU SER SER SER VAL MET ILE VAL SEQRES 10 B 256 LEU ILE ALA GLN ILE CYS GLY ILE ALA ASP ILE VAL ALA SEQRES 11 B 256 LEU LEU ALA ILE PHE GLY VAL ASN ALA SER MET ILE LEU SEQRES 12 B 256 PHE GLY TRP LEU GLN GLU LYS TYR THR GLN PRO LYS ASP SEQRES 13 B 256 GLY ASP LEU LEU PRO PHE TRP PHE GLY CYS ILE ALA GLY SEQRES 14 B 256 ILE VAL PRO TRP ILE GLY LEU LEU ILE TYR VAL ILE ALA SEQRES 15 B 256 PRO GLY SER THR SER ASP VAL ALA VAL PRO GLY PHE VAL SEQRES 16 B 256 TYR GLY ILE ILE ILE SER LEU PHE LEU PHE PHE ASN SER SEQRES 17 B 256 PHE ALA LEU VAL GLN TYR LEU GLN TYR LYS GLY LYS GLY SEQRES 18 B 256 LYS TRP SER ASN TYR LEU ARG GLY GLU ARG ALA TYR ILE SEQRES 19 B 256 VAL LEU SER LEU VAL ALA LYS SER ALA LEU ALA TRP GLN SEQRES 20 B 256 ILE PHE SER GLY THR LEU ILE PRO ALA SEQRES 1 M 122 MET VAL GLN LEU VAL GLU SER GLY GLY GLY LEU VAL GLN SEQRES 2 M 122 ALA GLY GLY SER LEU ARG LEU SER CYS ALA ALA SER GLY SEQRES 3 M 122 MET ALA PHE SER ILE SER ARG MET GLU TRP HIS ARG GLN SEQRES 4 M 122 ALA PRO GLY ASN GLU ARG GLU LEU VAL ALA ARG ILE THR SEQRES 5 M 122 GLY GLY GLY ARG THR THR TYR GLY ASP SER VAL LYS GLY SEQRES 6 M 122 ARG PHE THR ILE SER ARG ASP ASN ALA LYS SER MET VAL SEQRES 7 M 122 TYR LEU GLN MET ASN SER LEU LYS PRO GLU ASP THR ALA SEQRES 8 M 122 VAL TYR TYR CYS ASN THR GLY TYR PHE TRP GLY GLN GLY SEQRES 9 M 122 THR GLN VAL THR VAL SER SER ALA ALA ALA LEU GLU HIS SEQRES 10 M 122 HIS HIS HIS HIS HIS SEQRES 1 N 122 MET VAL GLN LEU VAL GLU SER GLY GLY GLY LEU VAL GLN SEQRES 2 N 122 ALA GLY GLY SER LEU ARG LEU SER CYS ALA ALA SER GLY SEQRES 3 N 122 MET ALA PHE SER ILE SER ARG MET GLU TRP HIS ARG GLN SEQRES 4 N 122 ALA PRO GLY ASN GLU ARG GLU LEU VAL ALA ARG ILE THR SEQRES 5 N 122 GLY GLY GLY ARG THR THR TYR GLY ASP SER VAL LYS GLY SEQRES 6 N 122 ARG PHE THR ILE SER ARG ASP ASN ALA LYS SER MET VAL SEQRES 7 N 122 TYR LEU GLN MET ASN SER LEU LYS PRO GLU ASP THR ALA SEQRES 8 N 122 VAL TYR TYR CYS ASN THR GLY TYR PHE TRP GLY GLN GLY SEQRES 9 N 122 THR GLN VAL THR VAL SER SER ALA ALA ALA LEU GLU HIS SEQRES 10 N 122 HIS HIS HIS HIS HIS HET RET A 301 20 HET ACT A 302 4 HET PX2 A 303 36 HET PLM A 304 18 HET PLM A 305 18 HET RET B 301 20 HET ACT B 302 4 HET PLM B 303 18 HETNAM RET RETINAL HETNAM ACT ACETATE ION HETNAM PX2 1,2-DILAUROYL-SN-GLYCERO-3-PHOSPHATE HETNAM PLM PALMITIC ACID FORMUL 5 RET 2(C20 H28 O) FORMUL 6 ACT 2(C2 H3 O2 1-) FORMUL 7 PX2 C27 H52 O8 P 1- FORMUL 8 PLM 3(C16 H32 O2) HELIX 1 AA1 THR A 5 ALA A 34 1 30 HELIX 2 AA2 VAL A 65 SER A 84 1 20 HELIX 3 AA3 GLY A 86 MET A 97 1 12 HELIX 4 AA4 ILE A 102 GLY A 124 1 23 HELIX 5 AA5 ASP A 127 THR A 152 1 26 HELIX 6 AA6 LEU A 159 ALA A 168 1 10 HELIX 7 AA7 ILE A 170 ALA A 182 1 13 HELIX 8 AA8 PRO A 192 GLY A 219 1 28 HELIX 9 AA9 ASN A 225 SER A 250 1 26 HELIX 10 AB1 THR B 5 ALA B 34 1 30 HELIX 11 AB2 VAL B 65 SER B 84 1 20 HELIX 12 AB3 GLY B 86 MET B 97 1 12 HELIX 13 AB4 ILE B 102 GLY B 124 1 23 HELIX 14 AB5 ASP B 127 THR B 152 1 26 HELIX 15 AB6 LEU B 159 ALA B 168 1 10 HELIX 16 AB7 ILE B 170 ALA B 182 1 13 HELIX 17 AB8 PRO B 192 GLY B 219 1 28 HELIX 18 AB9 ASN B 225 SER B 250 1 26 SHEET 1 AA1 2 ALA A 38 GLY A 43 0 SHEET 2 AA1 2 VAL A 58 PRO A 64 -1 O THR A 63 N LEU A 39 SHEET 1 AA2 2 ALA B 38 GLY B 43 0 SHEET 2 AA2 2 VAL B 58 PRO B 64 -1 O THR B 63 N LEU B 39 SHEET 1 AA3 4 VAL M 5 SER M 7 0 SHEET 2 AA3 4 LEU M 18 ALA M 23 -1 O SER M 21 N SER M 7 SHEET 3 AA3 4 MET M 77 MET M 82 -1 O MET M 82 N LEU M 18 SHEET 4 AA3 4 PHE M 67 ASP M 72 -1 N THR M 68 O GLN M 81 SHEET 1 AA4 6 GLY M 10 VAL M 12 0 SHEET 2 AA4 6 THR M 105 VAL M 109 1 O THR M 108 N GLY M 10 SHEET 3 AA4 6 ALA M 91 ASN M 96 -1 N ALA M 91 O VAL M 107 SHEET 4 AA4 6 GLU M 35 GLN M 39 -1 N GLU M 35 O ASN M 96 SHEET 5 AA4 6 ALA M 49 ARG M 50 -1 O ALA M 49 N TRP M 36 SHEET 6 AA4 6 THR M 58 TYR M 59 -1 O THR M 58 N ARG M 50 SHEET 1 AA5 4 VAL N 5 SER N 7 0 SHEET 2 AA5 4 LEU N 18 ALA N 23 -1 O ALA N 23 N VAL N 5 SHEET 3 AA5 4 MET N 77 MET N 82 -1 O MET N 82 N LEU N 18 SHEET 4 AA5 4 PHE N 67 ASP N 72 -1 N THR N 68 O GLN N 81 SHEET 1 AA6 6 GLY N 10 VAL N 12 0 SHEET 2 AA6 6 THR N 105 VAL N 109 1 O THR N 108 N GLY N 10 SHEET 3 AA6 6 ALA N 91 ASN N 96 -1 N ALA N 91 O VAL N 107 SHEET 4 AA6 6 GLU N 35 GLN N 39 -1 N GLU N 35 O ASN N 96 SHEET 5 AA6 6 ALA N 49 ARG N 50 -1 O ALA N 49 N TRP N 36 SHEET 6 AA6 6 THR N 58 TYR N 59 -1 O THR N 58 N ARG N 50 SSBOND 1 CYS M 22 CYS M 95 1555 1555 2.04 SSBOND 2 CYS N 22 CYS N 95 1555 1555 2.04 LINK NZ LYS A 241 C15 RET A 301 1555 1555 1.43 LINK NZ LYS B 241 C15 RET B 301 1555 1555 1.43 CRYST1 1.000 1.000 1.000 90.00 90.00 90.00 P 1 ORIGX1 1.000000 0.000000 0.000000 0.00000 ORIGX2 0.000000 1.000000 0.000000 0.00000 ORIGX3 0.000000 0.000000 1.000000 0.00000 SCALE1 1.000000 0.000000 0.000000 0.00000 SCALE2 0.000000 1.000000 0.000000 0.00000 SCALE3 0.000000 0.000000 1.000000 0.00000